Hi, I have been using reactive NHS fluorescent reagents and they lose activity after the vial is opened and kept as powder at 4oC. Dissolving them in DMSO and freezing them at -20oC also results in loss of reactivity.
Does anybody know of a way to store reactive NHS reagents in liquid form without losing reactivity towards amines?
We use a lot of NHS chemistry, and have working solutions that are 5 years old.
First, you have to make sure that you use water-free solvent to dissolve the NHS-ester. Dry DMF, stored under argon and over molecular sieves, works well for us.
Second, you have to make sure that the vial can equilibrate to room temperature before opening it. Otherwise you will have a lot of water condensing inside the vial, hydrolysing the NHS ester.
Third, when you plan to often use small amounts of the NHS ester, better aliquot the stock solution.
Using these simple precautions, our NHS esters usually retain their reactivity for a long time.
Storing at -20oC in the presence of dessicants in a well enclosed container.
Hi Andrei, I sent a reprint request to the authors of that paper, thanks for the tip!.
@Dmitri, I'm not sure I understand. What liquid are you suggesting that I store the NHS esters in, and what dessicants are you reccomending? Thanks.
If you really want them to preserve fluorescent NHS esters, handle them under a nitrogen atmosphere using a globe box in low ambient light or refill the container with nitrogen after opening. I'd recommend using small stock aliquots in anhydrous DMF/DMSO for each experiment but keep the bulk as the solid. Store them under nitrogen in the dark (cover the bottle in foil and/or use an amber glass bottle) in a freezer and keep the bottle in a zip-lock back that contains a desiccant. Make sure your DMSO is anhydrous. DMSO is very hydroscopic. If the bottle of DMSO doesn't have a rubber seal (like a sure seal) and used under nitrogen then I can guarantee it is loaded with water.
These esters hydrolyse very easily with water. First of all make sure the solvent you use is waterless. This is a different quality you can order these bottles as such or dry the solvent yourselve. As mentioned by Philip DMSO is very hygroscopic. I used DMF of analytical quality from a sealed container. Use a dried glass syringe and dried glassware to transfer the solvent from the container to the recipient, preferably in a glove box. Use brown glass, or alternatively pack the glass recipient with Al foil in order to avoid exposure to light. store in a fridge or freezer should be sufficient. If the quality of the freezer or fridge is poor (a lot of ice for example) your can take a plastic bottle, put some CaCl2 on the bottom, cover this with cottonwool, put the little recipient with your NHS-ester solution on top of that and seal this container. As such you create a small waterless environment. While handling these NHS-ester bare in mind to avoid exposure to light and water, including water from the air. Good luck !
We use a lot of NHS chemistry, and have working solutions that are 5 years old.
First, you have to make sure that you use water-free solvent to dissolve the NHS-ester. Dry DMF, stored under argon and over molecular sieves, works well for us.
Second, you have to make sure that the vial can equilibrate to room temperature before opening it. Otherwise you will have a lot of water condensing inside the vial, hydrolysing the NHS ester.
Third, when you plan to often use small amounts of the NHS ester, better aliquot the stock solution.
Using these simple precautions, our NHS esters usually retain their reactivity for a long time.
@Andrei, thanks for the reprint! The authors stress that dioxane they use is anhydrous.
@Phillip, Tim and Dominik. Thanks a lot for your expert answers. I really appreciate it!
For me, the take home lesson is that whatever solvents I use to dissolve NHS esters, they have to be anhydrous if I want to retain activity. Also, NHS esters have to be stored under anhydrous gasses.
Can you guys tell me if it matters whether I store NHS ester liquid aliquots at +4oC, -20oC or -80oC
Thanks.
We store our NHS esters - dissolved in DMF - at -20 °C. DMF has the advantage that it doesn't freeze at -20°C, so the solution doesn's go through repeated melting and thawing cycles.
When you have easy access to a -80°C freezer and have the time to make small aliquots of the NHS ester solution, then -80°C is for sure the better choice. But in anhydrous solvents, -20°C should be enough.
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